Noninvasive Investigation of Phloem Structure by 3D Synchrotron X-Ray Microtomography.

X-ray microtomography (µCT) is a three-dimensional imaging technique, which has, over the past decade, established itself as a go-to method for nondestructive visualization of plant tissue with submicrometer resolution. µCT is closely related to medical computed tomography, in that a measurement consists of acquiring a series of radiographs from different directions around the sample. Especially with synchrotron X-ray sources, these radiographs exhibit significant phase contrast. This greatly enhances soft tissue contrast, making it well suited for plant imaging. Tomographic reconstruction techniques are then employed to convert the stack of radiographs into a 3D volumetric image. Compared with the laboratory X-ray tube-based systems, synchrotron tomography beamlines also offer high throughput, with tens of samples scanned over the course of a typical 24-h beam time.Synchrotrons are typically operated as user facilities, with a staff member assisting users in aligning the beamline and all instrumentation-related matters. From the user's point of view, success of a synchrotron µCT experiment is often dependent on secure sample mounting, choice of appropriate beam parameters, and post-processing the data, i.e., extracting scientifically meaningful results from the 3D image. In this chapter, we review the issues to consider in preparation of a µCT experiment from the point of view of a phloem researcher, emphasizing those aspects which are directly under the user's control rather than technical specifics, which vary from one beamline to another.

The hydraulic architecture of Ginkgo leaves.

PREMISE OF THE STUDY: The hydraulics of xylem has been widely studied in numerous species and organ types. However, comparatively little is known about how phloem and xylem are hydraulically coupled or about many of the basic structural properties of phloem (such as conducting cell numbers and conductive areas), which nevertheless have direct bearing on understanding phloem loading and unloading. METHODS: Using a combination of light, epifluorescence, confocal, and transmission electron microscopy, we quantified the hydraulic architecture of Ginkgo biloba leaf laminae and examined the scaling relationships between phloem and xylem in five fully mature leaves. KEY RESULTS: The conductive areas and lengths of sieve cells and tracheids increase basipetally toward the petiole in a manner that is consistent with Münch's pressure flow hypothesis for phloem transport. This trend holds true for individual veins, the sum of conductive areas across all veins at any distance from the petiole, and for individual sieve cells and tracheids. Further, the conductive areas of phloem and xylem are isometrically correlated across the entire vasculature of the leaf lamina. The data for conducting cell areas do not conform with the predictions of the hydraulic models of da Vinci and Murray. CONCLUSIONS: The scaling of Ginkgo lamina hydraulics complies with that observed in leaves of other gymnosperms and most angiosperms and is inconsistent with theoretical models that assume that the volume of transported incompressible fluids is conserved.

Comparison of phloem and xylem hydraulic architecture in Picea abies stems.

The hydraulic properties of xylem and phloem differ but the magnitude and functional consequences of the differences are not well understood. Phloem and xylem functional areas, hydraulic conduit diameters and conduit frequency along the stems of Picea abies trees were measured and expressed as allometric functions of stem diameter and distance from stem apex. Conductivities of phloem and xylem were estimated from these scaling relations. Compared with xylem, phloem conduits were smaller and occupied a slightly larger fraction of conducting tissue area. Ten times more xylem than phloem was annually produced along the stem. Scaling of the conduit diameters and cross-sectional areas with stem diameter were very similar in phloem and xylem. Phloem and xylem conduits scaled also similarly with distance from stem apex; widening downwards from the tree top, and reaching a plateau near the base of the living crown. Phloem conductivity was estimated to scale similarly to the conductivity of the outermost xylem ring, with the ratio of phloem to xylem conductivity being c. 2%. However, xylem conductivity was estimated to increase more than phloem conductivity with increasing tree dimensions as a result of accumulation of xylem sapwood. Phloem partly compensated for its smaller conducting area and narrower conduits by having a slightly higher conduit frequency.